一株产乙醇嗜热厌氧油藏微生物Thermoanaerobacter sp DF3的性质研究

摘要

[目的]了解油藏微生物Thermoanaerobacter sp DF3 的生理生化特性,优化木糖产乙醇培养方案.[方法丁利用厌氧分离技术从大港油田油层采出液中分离到一株产乙醇厌氧杆菌DF3.采用生理生化鉴定与16S rDNA序列的系统发育学分析其系统发育地位,用气相色谱分析其代谢产物.[结果]菌株DF3是一株严格厌氧的嗜热细菌,呈直杆状,G-,菌体大小为0.42 μm×(1.60～5.20)μm,单生、成对或成串生长,产顶端芽孢;生长温度为45～78 ℃(最适65 ℃),能利用葡萄糖、木糖、果糖、核糖、甘露糖、阿拉伯糖、蔗糖、半乳糖、乳糖、纤维二糖、松三糖、棉子糖、淀粉等作为底物;其16S rRNA与T.pseudoethanolicus相似性为99.7%.发酵葡萄糖与木糖的主要产物为乙醇,培养方案优化后其代谢木糖产乙醇终浓度为2.0 g/L.[结论]通过试验证明菌株DF3是目前已知菌株中产乙醇活性较强的菌株之一,在65℃时代谢木糖能产生2.0g/L的乙醇.目前代谢木糖高产乙醇的菌株均由国外分离获得,菌株DF3的分离获得为我国研究木质纤维素产乙醇提供了优良的出发菌株.%[ Objective ] The research aimed to learn about the physiological and biochemical characteristics of the microbe Thermoanaerobacter sp DF3 in oil pool and optimize the culture plan of producing ethanol from xylose. [ Method ] A strain of anaerobic ethanol-producing bacillus DF3 was isolated from produced liquid from oil layer of Dagang oil field using anaerobic isolation technics. Physiological and biochemical identification and phylogenetic analysis of 16S rDNA sequence were used to analyze its phylogenetic position. Gas chromatographic analysis was used to analyze its metabolites. [ Result] The strain DF3 was one strict anaerobic thermophilic bacterium. It was straight in rod shape,Gram negative and its thallus size was 0.42 μm × ( 1.60 - 5.20 ) μm. It was solitary, in pairs or string and it produced apical spore. Its growth temperature was45-78 ℃ and 65 ℃ was optimum. All of glucose, xylose, fructose, ribose, mannose, arabinose, sucrose, galactose, lactose, cellobiose, melizitose, raffinose and starch could be used as substrate. The similarity of 16S rDNA between strain DF3 and T.pseudoethanolicus was 99.7％. The main product from glucose and xylose fermentation was ethanol. After the culture plan was optimized, the final ethanol conch. of xylose fermentation with DF3 was 2.0 g/L. [ Conclusion] It was proved through experiments that the strain DF3 was one of the strains with higher ethanol-producing activity among known ethanol producers at present and it could produce 2.0 g/L ethanol from xylose metabolization at 65 ℃. Now, all the high-yield ethanol producers of xylose metabolization were isolated by other nations. So, the isolation of the strain DF3 supplied a good initial strain for researching alcohol production from lignocellulose in China.